Apoptosis is a physiologically important process in the nervous system. Recent evidence indicates that apoptosis may also play a role in neuronal death in Alzheimer's disease and other neurodegenerative disorders. In apoptosis, profound alterations in the cytoskeleton occur which contribute to the characteristic morphological features, and may be due in part to the inappropriate phosphorylation of certain proteins. The microtubule-associated protein tau is a critically important phosphorylated cytoskeletal protein necessary for the maintenance of neuronal structure and function. A hallmark of Alzheimer's disease is the extensive site-specific hyperphosphorylation of tau, which is thought to be due to an imbalance in the activities of specific protein kinases and phosphatases. The relationships between apoptosis and tau hyperphosphorylation have not been studied previously. Preliminary studies from this lab indicate that in apoptotic PC12 cells expressing a neuronal phenotype, there are imbalances in specific protein kinases and phosphatases as well. In addition, preliminary findings indicate that Ser 202 in tau is hyperphosphorylated in differentiated PC12 cells undergoing apoptosis; a site that is also hyperphosphorylated in tau from Alzheimer's disease brain. Therefore the applicant's overall hypothesis is that tau is hyperphosphorylated at specific sites during apoptosis, and this hyperphosphorylation results in alterations in the metabolism and function of tau. Elucidating the mechanisms which contribute to the inappropriate hyperphosphorylation of certain sites on tau during apoptosis, and the functional consequences of these modifications should increase our understanding of the processes which result in the hyperphosphorylation of tau in Alzheimer's disease. The studies described in this proposal will use two models of apoptosis. The first is a well characterized model using differentiated PC12 cells deprived of nerve growth factor and serum, and the other model uses human neuroblastoma cells treated with ceramide in serum-free conditions. The goals of this proposal are to test the following hypotheses: (1) during apoptosis tau becomes hyperphosphorylated in a site-specific manner that progresses in parallel with the cell death process, and several of the sites on tau that are phosphorylated during apoptosis correspond to sites that are hyperphosphorylated on tau from Alzheimer's disease brain, (2) activation of specific protein kinases contribute to the site-specific hyperphosphorylation of tau during apoptosis, (3) the significant decrease in protein phosphatase 2B during apoptosis contributes to tau hyperphosphorylation and plays a facilitating role in the process, and (4) the hyperphosphorylation of tau which occurs during apoptosis results in functional alterations.